Anatomy defines the `reference' atlas for all of neuroscience. It is one of the most important markers of disease or damage to the brain, and constrains the circuitry of neural computation. However, brain maps are fundamentally incomplete. There remains an information and resolution gap at the mesoscale: whole brain maps visualized at micrometer resolution so that cell shapes, numbers, and positions along with their long range projections can be visualized in a single brain. Such maps, especially if they were compatible with the other modalities, could provide valuable information for determining the cellular composition of brains and serve as a vital bridge between studies using disparate resolutions (e.g. functional magnetic resonance imaging or MRI, and serial section electron microscopy) to image the brain. X-ray microtomography is the only approach that can provide mesoscale detail on whole brains without the need for slicing. We propose to use the nation-leading capabilities of the Advanced Photon Source (APS) at Argonne National Laboratory, which offers a source brightness millions of times higher than laboratory sources. With this source, we have already demonstrated that propagation-based monochromatic phase contrast x-ray imaging can be used to obtain high contrast tomograms of millimeter-sized regions of plastic embedded and metal-stained mouse brain, with data collection times of about a few minutes and a voxel resolution of one micrometer. We will develop whole brain sample preparation methods optimized for x-ray microtomography, and for correlative studies with serial section electron microscopy for synapse-level resolution of small regions (Aim 1). We will develop mosaic x-ray tomography to move from millimeter sized samples to whole mouse brains, with a path towards future studies of human brains (Aim 2). We will develop a high speed tomographic reconstruction workflow, and methods for volume segmentation, analysis, and visualization to make sense of these huge 3D datasets (Aim 3). Synchrotron-based x-ray microtomography has been unknown to most neuroscientists. It fills the information and resolution gap between MRI studies of living animals and humans, light microscopy of specific molecule types within a few millimeters of the brain surface, and electron microscopy studies with exquisite anatomical detail of limited regions. Our team includes experts in brain tissue preparation and electron microscopy, x-ray microtomography, and analysis of brain anatomy. We are therefore in a unique position to develop x-ray tomography for massive scale brain anatomy, and to make these advances available to the neuroscience community since the APS is a no-cost user facility.